Seroprevalence of Toxoplasma gondii in Females Aged 15–45 Years from Bihor County,Western Romania

Introduction

Toxoplasmosis is a common infectious disease caused by a single-celled protozoan parasite named Toxoplasma gondii (T. gondii) (Montoya and Liesenfeld 2004). Transmission of this parasite to humans occurs through the ingestion of food and water contaminated with oocysts shed by infected cats in the acute phase of the infection (Elmore et al. 2010, Lilly et al. 2013). Another route of transmission to humans is through the consumption of raw or undercooked meat that contains tissue cysts (Dubey 2004, Dawson 2005).

Although the acute disease passes asymptomatically in most immunocompetent patients, clinical signs may be present particularly in immunocompromised patients and congenitally infected children (Montoya and Liesenfeld 2004, Olariu et al. 2019). If infection is acquired during or shortly prior pregnancy, the offspring may be severely affected due to the clinical manifestations that may occur, including mental retardation, seizures, blindness, and even death (Kravetz and Federman 2005, Remington and Klein 2016). Therefore, the seroprevalence of T. gondii antibodies should be monitored in females of reproductive age (Olariu et al. 2019).

The global seroprevalence of IgG antibodies against T. gondii in pregnant females is 32.9%, South America presenting the highest seroprevalence (45.2%) and Western Pacific the lowest (11.2%) (Bigna et al. 2020). The prevalence of IgG antibodies in females aged 15–45 years located in Europe varies from country to country: 34.7% in Turkey, 22.3% in Poland, and 39.5% in Germany (Warnecke et al. 2020). A higher T. gondii seroprevalence was associated with low income and developing countries (Rostami et al. 2020).

Bihor County is one of the largest counties located in Western Romania alongside Arad County and Timis County, two well-known endemic areas (Mihu et al. 2020, Olariu et al. 2015, Olariu et al. 2020). Epidemiological information on the prevalence of T. gondii infection in Romania is incomplete and no national program for screening of pregnant females is currently implemented. Moreover, data on T. gondii seroprevalence in females aged 15–45 years are limited and scant. Currently, T. gondii seroprevalence is unknown in Bihor County, Romania. Therefore, we assessed the seroprevalence of T. gondii antibodies in females aged 15–45 years located in this region.

Materials and Methods

In this study, 1935 females aged 15–45 years were tested for T. gondii IgG antibodies. No clinical criteria were used to include the females in the study. The participants were aged between 15–45 years and were residents of Bihor County, with a population of 575,398 inhabitants. Samples included in this study were collected from January 01, 2016 to December 31, 2018.

Blood samples were collected using standard venipuncture methods into Serum Separation Gel & Clot Activator Vacuum Tubes. The collected samples were centrifuged 4000 g for 10 min, 10–30 min after collection. Sera were analyzed for IgG antibodies against T. gondii within 2 h of the initial blood draw.

IgG antibodies against T. gondii were determined using ARCHITECT I2000SR (Abbott Diagnostics Total Solution, Germany) in accordance with the manufacturer's instructions and internal laboratory standards. This assay uses a two-step chemiluminescent microparticle immunoassay to quantitatively determine the specific IgG antibodies to T. gondii. Specimens with concentration values ≥3.0 IU/mL were considered reactive, concentration values from 1.6 to 2.9 IU/mL equivocal, and concentration values <1.6 IU/mL were considered nonreactive for IgG antibodies against T. gondii. For the purpose of this study equivocal results were considered negative.

The females included in this study were grouped into six categories based on their age when the sample was drawn: 15–19 years, 20–24 years, 25–29 years, 30–34 years, 35–39 years, and 40–45 years.

Urban areas in this study were defined as places with a high population density and refers to a city, town, or suburb. Rural areas were defined as places with a lower density of population located outside cities or towns and refers to a village or any human settlement not included within an urban area.

Data were collected using Microsoft Excel, version 2011 (Microsoft Corp., Redmond, WA). Statistical analyses were performed with the Epi Info statistical package 3.3.2 (Centers for Disease Control and Prevention, Atlanta, GA). Mantel–Haenszel test was used for comparison between groups. Logistic regression was used to examine the association between positive cases, areas of residence, and age. Crude odds ratios and their 95% confidence intervals (95% CIs) were calculated. The logistic regression was conducted with Stata 16.1 (StataCorp). Statistical significance was set at p < 0.05. This study was approved by the Victor Babes University Ethics Committee, Timisoara, Romania (no.2 from 08.01.2018).

Results

IgG antibodies against T. gondii were found in 706 of 1935 (36.48%) females aged 15–45 years and the proportion of IgG positive females tended to increase with age. No statistically significant differences in seroprevalence were observed when comparing females aged between 15 and 19 years with those between 20 and 24 years (73 out of 198, 36.87%, p = 0.21), 25–29 years (265/741 [35.76%], p = 0.17), 30–34 years (239/624 [38.3%], p = 0.12), 35–39 years (97/286 [33.92%], p = 0.3), and 40–45 years (25/56 [44.64%], p = 0.63) (Table 1).

If area of residence was taken into account, females residing in rural areas had a statistically significant higher seroprevalence of IgG T. gondii antibodies when compared with females residing in urban regions (304 out of 636 vs. 402 out of 1293, p = 0.001). When data were further stratified by both age and area of residence, a statistically significant seroprevalence difference was found in females residing in rural areas aged between 20 and 24 years (20 out of 91 vs. 53 out of 107, p = 0.038), 25–29 years (141 out of 467 vs. 124 out of 274, p < 0.001), 30–34 years (154 out of 454 vs. 85 out of 170, p < 0.001), and 35–39 years old (66 out of 229 vs. 31 out of 57, p < 0.001) when compared with females in the same age groups from urban regions (Table 1).

The logistic regression analysis revealed that females residing in rural areas were twice as likely to be positive compared with females residing in urban areas. OR = 2.04 (p < 0.001; 95%CI: 1.68–2.48).

Discussions

This study revealed a T. gondii seroprevalence of 36.48% in females aged 15–45 years residing in Bihor County, Romania. This seroprevalence is higher than the seroprevalences reported in females from Poland (22.3%), lower than the prevalence reported in Germany (56.2%), and similar to those in Turkey (34.7%) (Warnecke et al. 2020).

Compared with the recent studies performed outside Europe, the seroprevalence of T. gondii IgG antibodies in our females aged 15–45 years was >33% prevalence reported in Iran (Mizani et al. 2017), 27.8% in Saudi Arabia (Alzaheb 2018), and 27% in Sudan (Mustafa et al. 2019), but it was <40% prevalence reported in Cameroon (Wam et al. 2016).

The seroprevalence of T. gondii IgG antibodies in females aged 15–45 years may vary between countries and sometimes between regions located within the same country (Fanigliulo et al. 2020). In Western Romania, for instance the seroprevalence in females aged 15–45 years was 51.9% in Timis County (Capraru et al. 2016) and 41.16% in Arad County (Mihu et al. 2020). We cannot explain the difference in seroprevalence between these counties, because we did not have additional information regarding potential risk factors (meat consumption, contact with soil or pets, educational level, and profession) associated with T. gondii infection in our participants and no questionnaire was used in this study. It is likely that some of these exposure factors may contribute to the difference in seroprevalence between counties.

We found a significant higher T. gondii seroprevalence in females residing in rural areas compared with those from urban areas and these results are similar with those reported by former authors (Liu et al. 2009, Vilibic-Cavlek et al. 2011, Gao et al. 2012, Bigna et al. 2020). Our findings revealed that females residing in rural areas were twice as likely to be T. gondii positive compared with females residing in urban areas.

In rural areas, the communities are more likely to obtain water from an open source, own a cat, consume unwashed vegetables obtained from the community, and consume locally produced undercooked meat (Munoz-Zanzi et al. 2016). However, some studies (Aqeely et al. 2014, Morais et al. 2021) found women in urban areas to have higher seroprevalence of T. gondii antibodies when compared with females residing in rural areas. These findings were attributed to the different habits of urban residents as eating junk food and poultry from restaurants, which have been found to represent a source of transmission for T. gondii (Aqeely et al. 2014) or by the conditions of overpopulation and contamination of the water supply (Morais et al. 2021). To further understand this difference, studies that include questionnaires should be conducted in this region to evaluate additional risk factors for toxoplasmosis.

Limited data are currently available on the seroprevalence of T. gondii in animals from Romania. A study conducted by Györke and colleagues revealed that 47% of house cats had T. gondii antibodies (Györke et al. 2011). Darabus et al. (2011) and Dubey (2004) reported a T. gondii seroprevalence from 36.3% to 61.3% of the sheep population. T. gondii antibodies were also demonstrated in 57.7% of the backyard animals (Balea et al. 2012)

This is the first study that assessed the seroprevalence of T. gondii in females aged 15–45 years from Bihor County, Romania. Our findings may be a valuable starting point for counseling and education programs on toxoplasmosis and for the implementation of a screening program for pregnant females at the national level. This study also provides an insight into the demographic risk factors as a basis for a future prevention program for T. gondii infection and represents another piece in the jigsaw puzzle of the epidemiology of toxoplasmosis.